Oxidized fatty acids and their metabolites, termed oxylipins, act as signaling molecules in virtually all kingdoms of life . For the model ascomycete Aspergillus nidulans, it has been shown in recent years that specific oxylipins, known as precocious sexual inducers (psi factors) play a crucial role for the balance of different developmental processes and the formation of mycotoxins [2–4]. From a chemical point of view, these signaling molecules are fatty acids with 18 carbon atoms, which are hydroxylated at distinct positions. The finding that these oxylipins are involved in processes related to fungal pathogenicity and that there is a chemical similarity between the pathogen-derived signal molecules and host-derived oxylipins led to the proposal that they might play a central role in pathogen–host interaction and trans-species communication, the so-called cross-kingdom signaling [5–7]. After the genome of A. nidulans was sequenced, three genes putatively involved in formation of these important signaling molecules have been identified and were named ppoA, ppoB and ppoC . With the advent of more and more fungal genomes sequenced, it became obvious that the enzymes encoded by these and homologue genes are conserved in virtually all ascomycetes [1,8] and play a pivotal role in conferring virulence of ascomycetes towards mammalian and plant hosts [3,7,9]. These psi factor producing oxygenases (Ppos) are heme enzymes consisting of two distinct enzymatic functionalities . To date the enzyme encoded by A. nidulans ppoA, beside the 7,8-linoleate diol synthase (7,8-LDS) from Gaeumannomyces graminis , is by far the biochemically best characterized [10,12–14]. It was shown that PpoA is a bifunctional, tetrameric heme enzyme with a molecular mass of ∼ 480 kDa [10,12–14]. Both domains show functional similarities and sequence homologies to known oxylipin-forming enzymes from mammals and plants. In a first step the substrate, an unsaturated fatty acid with 18 carbons, is converted to a C8-hydroperoxy fatty acid at the N-terminal enzyme domain that is proposed to have high structural and functional similarity to the mammalian prostaglandin H2 synthase (PGHS). The C-terminal domain of the polypeptide chain is predicted to possess a conserved P450-fold and catalyzes the rearrangement of the N-terminally formed hydroperoxy fatty acid to a dihydroxy fatty acid. Based on the reaction catalyzed, we propose that the P450 domain of the fungal PpoA enzyme shares the highest similarity to the subfamily of class III P450s . This subclass comprises all P450s that do not require external electron donors and thus do not catalyze typical P450 monooxygenations, e.g. they do not insert oxygen, derived from molecular oxygen. The enzymes of this class instead utilize fatty acid peroxides as substrates for rearrangement reactions to yield diverse products. Interestingly, all known enzymes of this subclass, e.g. Cyp5 (thromboxane A synthase), Cyp8a (prostacyclin synthase, PGIS) and Cyp74 [allene oxide synthase (AOS), divinyl ether synthase and hydroperoxide lyase], are involved in the biosynthesis of oxylipins in various species .
While information on the mechanism of the two reaction steps, fatty acid dioxygenation and rearrangement of the formed hydroperoxy fatty acid, has accumulated through the last few years [10,12–14], structural information on the enzyme is still scarce. Therefore, in this work we predict the 3D fold of the single PpoA domains, independently. Based on these hypothetical models, we could validate the proposed conservation of oxylipin-forming enzymes and derive new determinants for fatty acid oxidation in the single domains of the prototype enzyme PpoA.